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Designing miRNA involves three steps. After specifying the input data, the miRArchitect engine performs calculations. The submitted sequence is then sent to the server, where the amiRNA design process begins. Once processing is complete, the user is redirected to the results page. Please be patient, as this may take some time.

1. Data upload

1.1. Sequence search

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Users can search for sequences using either the sequence name (e.g., "BRCA1") or the Ensembl identifier (e.g., "ENSG00000012048") to find all matching genes and transcripts. After clicking "Proceed," a table with detailed information will be displayed on the next page.

1.2 Custom sequence and/or target

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Users can input a custom sequence and/or target. Targets must be exactly 21bp in length and should be separated by a semicolon or by pressing Enter. Users can click "X" to delete a specific target.

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Each task is assigned a unique ID. By entering this number, the user is redirected to the results page. The link remains active for 48 hours.

2. Target specification

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3. Design parametrization

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• Min GC content [%] - Minimum GC percentage
• Max GC content [%] - Maximum GC percentage
• Max GC stretch - Maximum length of a consecutive subsequence made of G and C
• Bps to init binding - Bps to init binding, used when checking for off-targets
• Bps to init duplex dissociation - The number of nucleotides which are checked on both ends to determine if the duplex will start to dissociate from the correct side
• Min MFE difference between ends - Minimum energy difference in kcal/mol between dissociation energies of both ends of the duplex, one end has to have lower energy in order to be always dissociated first
• 2D struct. differences - Maximum number of differences between the result amiRNA 2D structure and the natural 2D structure
• Max melting temperature - Maximum melting temperature
• Different prefixes - Whether the guide strands with prefixes different from the original amiRNA should be considered
• Filter off-targets - Should the off-targets be filtered before designing the amiRNA?
• Invalid prefix penalty - Value added to the score if prefix is not identical with the original amiRNA
• amiRNA ID - ID of the template

4. Design and results